Inappropriate apoptosis is the cause of many human diseases, including cancer, neuronal degeneration, hearing loss, organ failure, myelodysplastic syndrome, multiple sclerosis, type 1 diabetes mellitus, thyroiditis, and ulcerative colitis. Compounds that affect apoptosis are potential therapeutics. Cell based assays are widely used in drug development, however, many promising candidates fail in subsequent mammalian testing. Zebrafish assays can serve as an intermediate step between cellular evaluation and mammalian testing. Zebrafish has several advantages, including transparent embryos for easy visual analysis, rapid embryonic development and low cost. Furthermore, similar pharmacological responses to those in the mammalian apoptotic machinery are present in zebrafish. Currently the standard whole animal apoptosis assay is TUNEL, a histochemical staining technique which detects apoptotic cells in sectioned tissue. This assay is time-consuming and permits examination at only a single time point. Since many degenerative diseases occur in stages, assay formats that permit continuous examination of changes in the apoptosis pattern and the duration of drug effects are needed. Using zebrafish embryos, this proposal will develop a quantitative microplate assay for high throughput screening of compounds. Complementary visual and flow cytometry based assays will also be developed to provide comprehensive analysis of apoptosis in zebrafish.
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| Parng, Chuenlei; Anderson, Nate; Ton, Christopher et al. (2004) Zebrafish apoptosis assays for drug discovery. Methods Cell Biol 76:75-85 |
